The central hypothesis of this proposal is that there is a subgroup of malignant human gliomas (MHG) in which the neoplastic cells gain a growth advantage over non-neoplastic cells by increased expression of the epidermal growth factor receptor (EGFR). EGFR expression will be determined in a large series of MHG biopsies, permanent MHG-derived cell lines and athymic mouse-grown MHG. Molecular and chromosomal analyses will define the mechanisms of increased EGFR expression and will determine if the amplified and/or rearranged genes are in double minute chromosomes, homogeneously staining regions, abnormally banded regions, or chromosomal translocations. Specific aims of this proposal are: (1) Antibodies against the EGFR will be used immunohistochemically on biopsied MHG, permanent MHG-derived cell lines, and MHG serially passaged in athymic mice to confirm that glioma cells express these receptors. Competition radioimmunoassay and epidermal growth factor (EGF) binding assays will be used to determine the range in level of receptor expression exhibited by these tumors, cell lines, and mouse-grown lines. 2) Immunoblot analysis, immunoprecipitation and autophosphorylation will be used to determine if the molecules detected in the immunologic studies are intact, functional EGFR or are EGFR-related proteins. 3) The mechanism of increased EGFR expression in the tumors, cell lines and mouse-grown tumors identified in Specific Aim (1) will be defined by assessing gene amplification, gene rearrangement, and gene transcription using a c-DNA probe against the EGFR in Southern and Northern blotting. 4) Since the panel of glioma biopsies, glioma-derived cell lines and athymic mouse-grown tumors which is available has been karyotyped, it will be possible to determine which of the gross chromosomal abnormalities, seen in these tumors, are associated with which mechanisms of increased expression. In situ hybridization will be performed on prophase and metaphase chromosomes from these specimens to confirm the location of amplified or translocated EGFR genes. If MHG cells differ from non-neoplastic cells by either quantitative or qualitative differences in EGFR expression, antibodies, polypeptides or other molecules which bind the EGFR could be used either alone or carrying toxic substances to retard tumor growth. Since the frequency and level of increased EGFR expression will be determined for MHG and the chromosomal and molecular mechanisms of enhancement of this gene product will be defined, the proposed studies will provide the theoretical basis for such therapeutic approaches. These experiments will also identify MHG cell lines and athymic mouse-grown tumors with high EGFR production which could be used for testing anti-EGFR agents in in vitro and in vivo systems.